Human skin is colonized by a diverse array of microorganisms. Colonization generally begins shortly after birth when an infant is exposed to the maternal microflora and other environmental events that typically lead to the colonization of a previously gnotobiotic human fetus. From the time of initial colonization, human skin remains in a state of flux where the composition of its resident microflora changes over time in response to factors intrinsic and extrinsic to the host.
In general, the microorganisms that colonize human skin may be grouped into three distinct categories: (1) those that are sporadic residents and typically do not proliferate on human skin, (2) those that may proliferate and remain on the skin for relatively short periods of time, and (3) those that may permanently colonize the skin. The members of these three groups may differ with respect to their preferred location on the skin and/or body of a person. Although human skin may be generalized as a cool, acidic, desiccate environment, a variety of microenvironments may be found in various locations on the surface of the skin. For example, the groin, axillary vault, and toe web typically exhibit higher temperature and humidity than other regions of the skin and/or body, which may promote the growth of microorganisms suited for such a microenvironment (e.g. Staphylococcus aureus and Corynebacteria). In another example, the sebaceous glands typically present on the face, chest, and back of a human may promote the growth of lipophilic microorganisms like Propionibacterium. Changes in diet; occupation; clothing choice; or the use of antibiotics, antibacterial soaps, moisturizers, cosmetics, hand sanitizers, and/or other anti-microbial skin products are also known to contribute to the variation observed in the type and/or amount of human skin microflora. Environmental factors like temperature, humidity, and exposure to ultra violet radiation are also known to cause changes in the type and/or amount of human skin microflora. Further, intrinsic host factors such as the host's genome, age, sex, and stage of sexual maturity may influence the state of the human skin microflora.
At least some members of the human skin microbiome provide benefits to their human host, for example, by stimulating the human immune system and/or producing anti-microbial substances. For example, Staphylococcus epidermidis has been shown to produce anti-microbial peptides that inhibit S. aureus biofilm formation. On the other hand, perturbations which disrupt the delicate balance of the skin microflora may result in undesirable consequences to the host and/or microflora. For example, increased production of free fatty acid byproducts associated with the proliferation of Propionibacterium acnes may promote the development of acne. Despite the diversity and/or fluctuations observed in the human microbiome among different individuals, it is believed that some members of the human microbiome may be common among different humans. In this regard, it has been shown that certain organisms typically constitute a significant portion of the human skin microbiome.
To combat any undesirable health and/or cosmetic consequences imposed on the host by the growth and/or activity of certain members of the skin microbiome, a variety of bactericidal agents (e.g., antibiotics) are known in the art. While the use of bactericidal agents may be clinically effective in reducing the symptoms associated with the growth of harmful microorganisms on human skin, there are drawbacks. For example, bactericidal agents such as topical antibiotics, benzoyl peroxide, and azelaic acid tend to affect both the beneficial and undesirable skin microflora indiscriminately. The death or behavioral change in the beneficial skin microflora in turn may lead to further undesirable health and/or cosmetic effects on the host, such as skin irritation. Moreover, certain bactericidal agents, in particular topical antibiotics, may promote antibiotic-resistant microbiota, sometimes referred to as “super bugs.”
A more advantageous strategy to combat any undesirable health and/or cosmetic consequence brought about by perturbations that disrupt the balance of the skin microbiota may be to identify agents that exhibit prebiotic activity for those members of the skin microbiome that produce a benefit to the host. Compositions containing such prebiotic agents could then be formulated by combining the prebiotic agent with an acceptable dermatological carrier and used topically. For example, moisturizers, hand and/or body soaps, cosmetics, hand sanitizers, body lotions, and/or other skin products suitable for human use may be formulated to include prebiotic agents. Skin care compositions that include a prebiotic agent may provide a more desirable alternative to conventional bactericidal agents, for example, by reducing the likelihood of skin irritation.
Currently, only a limited number of agents have been identified as exhibiting prebiotic activity on certain members of the human skin microbiome. There is no generally accepted method known in the art for effectively predicting which of the myriad of potential prebiotic agents will exhibit suitable prebiotic activity on skin microflora and be suitable for incorporation into topical skin care compositions. As a result, conventional methods for screening prebiotic agents may employ a difficult, time-consuming, and laborious battery of assays to identify a desired prebiotic agent. Additionally, the rich media used in conventional assays do not provide the desired sensitivity when attempting to detect prebiotic activity related to a particular test agent. In other words, a suitable prebiotic test agent may be overlooked due to the lack of sensitivity of conventional assays.
Those skilled in the art have long sought a suitable high-throughput screening method for identifying agents exhibiting prebiotic activity on members of the human skin microbiome, yet have been unsuccessful in developing such a method due to the variety of problems associated with its development. For example, the variability of the skin microbiota among individuals; expense of the assay; test volumes required for the assays; media choice; choice of cell types; detection sensitivity; difficulty in obtaining consistent data for small volumes of cultures; assay format; and the time required to conduct the assay individually and collectively contribute to the difficulty associated with the development of an industry-suitable, prebiotic high-throughput assay. Even identifying a suitable medium for such an assay is a laborious task due to the unique nutritional and environmental requirements of certain members of the human skin microbiome. In addition, there is desire to develop a tiered assaying methodology incorporating a high thru-put assay in combination with a low thru-put assay that is perhaps directionally more predicative for the commercial, large-scale screening of potential prebiotic compounds for the skin prior to placement of an expensive, time-consuming in vivo test. Accordingly, there is a need for a method of screening test agents that exhibit prebiotic activity on one or more of the skin commensal microorganisms of different individuals, and which uses a single, relatively simple media that provides suitable sensitivity. There is also a need for a high-throughput screening assay that identifies test agents exhibiting prebiotic activity on skin commensal microorganisms and that is relatively fast, inexpensive, and reliable.